About 5% of B cells in healthy mice and humans are allelically or isotypically included and hence co-express two different antibodies. In mice, dual antibody B cells (B_2R) expand with systemic autoimmunity, co-express autoreactive and non-autoreactive antibodies, and participate in immune responses, but this phenomenon is strain dependent. This study was developed with two goals: 1) to establish the contribution of TLR and IFN receptor signaling to the development of germinal center B cells that express two antibodies in MRL/lpr mice; and 2) to determine whether B_2R B cells are increased and particularly activated in a subset of adult patients diagnosed with systemic lupus erythematosus (SLE). Results from the MRL/lpr studies indicate that the enhanced differentiation of dual-κ B cells into germinal center B cells is due to a heightened response to TLR7 and TLR9 signaling, further fueled by an increased response to type II IFN. To understand the clinical and translational implications of our observations in mouse B_2R B cells, cohorts of SLE patients and healthy controls were recruited and evaluated for expression of dual BCRs. Results from flow cytometry and microscopy revealed supraphysiological frequencies of κ⁺λ⁺ B_2R cells in one fourth of the SLE patients. Abnormal numbers of κ⁺λ⁺ B cells correlated with higher frequencies of activated naïve B cells and age-associated B cells, and a lower proportion of "B cells that are naïve IgD⁺" (BND). However, results from single cell V(D)J sequencing demonstrated that these high κ⁺λ⁺ SLE patients harbored normal frequencies of κ⁺λ⁺ and other B_2R B cells. and we further show that their B cells were instead decorated by κ and λ VH4-34 autoantibodies. Thus, our findings indicate that elevated flow cytometric detection of isotypically-included B cells can identify patients with high titers of B cell-reactive VH4-34 autoantibodies and abnormal distribution of B cell subsets relevant to autoimmunity.

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